Basket Trials: Past, Present, and Future

Large-scale tumor molecular profiling has revealed that diverse cancer histologies are driven by common pathways with unifying biomarkers that can be exploited therapeutically. Disease-agnostic basket trials have been increasingly utilized to test biomarker-driven therapies across cancer types. These trials have led to drug approvals and improved the lives of patients while simultaneously advancing our understanding of cancer biology. This review focuses on the practicalities of implementing basket trials, with an emphasis on molecularly targeted trials. We examine the biologic subtleties of genomic biomarker and patient selection, discuss previous successes in drug development facilitated by basket trials, describe certain novel targets and drugs, and emphasize practical considerations for participant recruitment and study design. This review also highlights strategies for aiding patient access to basket trials. As basket trials become more common, steps to ensure equitable implementation of these studies will be critical for molecularly targeted drug development.


INTRODUCTION
Cancer has traditionally been classified and treated on the basis of a tumor's tissue of origin.Advances in genomic sequencing and large-scale efforts to molecularly profile tumors have identified recurrent genomic alterations across different cancer types, suggesting a unifying biology that may be susceptible to therapies targeting the aberrant pathway.As the number of potential targets has rapidly grown, there has been a shift toward a more diseaseagnostic, biomarker-driven approach to drug development.Basket trials enroll patients on the basis of a specific biomarker regardless of tumor histology.Due to their efficiency in testing biomarker-driven hypotheses, and facilitated by the increased availability and quicker turnaround times of comprehensive molecular profiling, basket trials have rapidly become the study design of choice for early-phase targeted therapy trials (Adashek et al. 2023, Bedard et al. 2020, Cunanan et al. 2017, Donoghue et al. 2020, Haslam et al. 2023, Hyman et al. 2017b, Murciano-Goroff et al. 2020a, Tao et al. 2018, Tsimberidou et al. 2020, Wahida et al. 2023, West 2017).
Basket trials have led to many notable successes.In May 2017, the US Food and Drug Administration (FDA) issued its first tissue-agnostic regulatory approval: Pembrolizumab, a PD-1 inhibitor, was approved for adult and pediatric patients with unresectable or metastatic microsatellite instability-high (MSI-H) or mismatch repair-deficient (MMRd) solid tumors (André et al. 2020, Blumenthal & Pazdur 2018, Boyiadzis et al. 2018, Goldberg et al. 2018, Lemery et al. 2017).Soon thereafter, in November 2018, the FDA approved larotrectinib, a small-molecule inhibitor of TRK proteins, in adult or pediatric patients harboring NTRK fusions (Drilon 2019, Drilon et al. 2018).The pivotal studies leading to these landmark approvals were remarkable for their clear demonstration of efficacy regardless of tumor type.Several additional tumor-agnostic approvals for advanced solid tumors have followed (Duke et al. 2022, Gouda et al. 2023, Lavacchi et al. 2020, Moore & Guinigundo 2023, Tateo et al. 2023) (Figure 1).
Basket trials allow for interrogation of genomic alterations that are too rare to be practically studied in a disease-specific manner.Additionally, they provide a way to investigate rare tumor types for which independent trials would not be feasible (Tao et al. 2018).For example, NTRK fusions are enriched in infantile fibrosarcoma, secretory breast carcinoma, and mammary analog secretory carcinomas, diseases whose treatment has been revolutionized by the tissue-agnostic approval of larotrectinib (Siozopoulou et al. 2021).
The potential of basket trials lies not only in the opportunity for tumor-agnostic drug development but also in the wealth of knowledge generated by including multiple histologies and, in some cases, multiple biomarkers within the same trial (Murciano-Goroff et al. 2020a).The scientific insights gained from these studies improve our understanding of cancer biology, which can ultimately be exploited to develop more precise biomarkers of sensitivity and resistance to therapy, improve patient selection for trials, and create better drugs (Murciano-Goroff et al. 2020a, Rosen et al. 2022, Tao et al. 2018, Vasan et al. 2019).The prevalence of basket trials as a tool for accelerating drug development demands reflection on how the design of these trials can be optimized and tailored to address varied targets, heterogeneous drug classes, and diverse patient populations.In the remainder of this review, we discuss the practicalities of implementing basket trials based on tumor sequencing.

OPTIMIZING BIOMARKER SELECTION
Improvements in genomic sequencing technologies have fueled the proliferation of basket trials.Prior to 2005, clinicians commonly used single-analyte tests relying on a polymerase chain reaction with Sanger sequencing to interrogate one mutation at a time.Consequently, tumors were typically tested only for biomarkers that had already been validated for a specific histology (Karlovich & Williams 2019).Over the past two decades, massive parallel DNA sequencing capable of evaluating multiple genes simultaneously has become widely available (Cronin & Ross 2011).This next-generation sequencing (NGS) approach continues to become faster, less expensive, more sensitive, and more accessible.Commercial NGS panels often contain more than 500 cancer-associated genes.These panels can detect mutations, insertions, deletions, copy number alterations, and some structural alterations (Morganti et al. 2019, Syn et al. 2016).Although not yet routinely used, whole-exome sequencing and whole-genome sequencing are becoming more widely available (Morganti et al. 2019).Improved detection of alterations has increased opportunities for therapeutic intervention.
In parallel, the number of therapies designed for specific genomic alterations in patients' tumors has risen (Berger & Mardis 2018).Matching of patients to molecularly selected therapy has historically been considered dichotomous, with a tumor either having or not having the alteration of interest.However, the breadth of information provided by NGS testing has highlighted the molecular complexity of tumors, posing new challenges in addition to opportunities for identifying better biomarkers for novel treatments (Murciano-Goroff et al. 2020a).
The key question when presented with an NGS report is whether the tumor contains a biomarker that may confer sensitivity to an approved or investigational therapy.Interpretation can be difficult, given the multiple variables that may influence tumor biology.Both histologic context and specific genetic/proteomic features (such as the unique amino acid change within a mutated gene, the level of amplification, or the coalterations observed) must be considered (Murciano-Goroff et al. 2020a).Ideally, genomic predictors of efficacy would be determined prior to designing trials, enabling enrichment for those subjects with the greatest chance of benefit.Practically, however, limitations on the scalability of preclinical modeling as well as the diversity of tumor histologies and molecular alterations have translated into bidirectional scientific investigations.As our understanding of cancer biology improves, we become better equipped to design rational biomarker-driven basket trials that enrich for those patients most likely to benefit.The results of clinical trials, in turn, advance biological knowledge, creating a positive cycle that has led to recent exponential growth in oncologic drug development (Bedard et al. 2020;Donoghue et al. 2020;Hyman et al. 2017b;Murciano-Goroff et al. 2020a,b) (Figure 2).

Identifying Driver Alterations
Tumors typically contain numerous genomic alterations.Importantly, not all alterations have an equal role in tumorigenesis and tumor maintenance; there are different biologic consequences depending on the alteration (Chang et al. 2018, Li et al. 2023, Torkamani & Schork 2008).When identifying patients for a basket trial, it is important to consider whether the alteration being targeted is critical for the cancer's survival.Preclinical modeling, including in vitro and in vivo studies, remains a useful tool for investigating whether a given alteration is oncogenic and likely to respond to a drug (Federici & Soddu 2020, Murciano-Goroff et al. 2020a).Unfortunately, it is not feasible to test every genomic alteration in the laboratory, and results may depend on the molecular and histologic context of the models used.
Computational analyses of large genomic data sets have identified recurrent mutations that occur in cancer more often than would be predicted by chance alone.These hot spots are often a consequence of positive selection, suggesting that they are important for tumor growth and/or survival and may be susceptible to targeted therapy (Chang et al. 2018).For example, this strategy identified small in-frame duplications in AKT as functionally relevant, and subsequent preclinical and clinical data confirmed that these alterations confer sensitivity to AKT inhibition (Shrestha Bhattarai et al. 2022).Other techniques to predict the biologic importance of variants of unknown significance (VUSes) include protein function prediction (such as using the PolyPhen-2 or SIFT tool) (Flanagan et al. 2010, Kerr et al. 2017), evolutionary conservation analysis (such as using GERP, CADD, or MutationAssessor) (Dong et al. 2015), saturation mutagenesis (Findlay et al. 2014), and multiplexed assays (Gasperini et al. 2016).
Notably, true driver oncogenes are often mutually exclusive in the treatment-naïve setting.In lung cancer, for example, de novo activating alterations in EGFR, KRAS, ROS1, ALK, RET, and NTRK rarely coexist (Farago & Azzoli 2017).The co-occurrence of EGFR and KRAS mutations negatively affects lung cancer development by inducing oncogenic stress and leading to synthetic lethality (Unni et al. 2015).The co-occurrence of a VUS alongside a known driver raises suspicion that the VUS is not itself a driver, though there are important exceptions, which may be disease dependent.In endometrial cancer, for example, multiple concurrent mutations involving PIK3CA and PTEN frequently coexist, amplifying PI3K/AKT pathway signaling (Sivakumar et al. 2023).Specific features of a genomic alteration may also provide clues as to whether it is likely to be therapeutically relevant.Examples include the level of gene amplification (Lai et al. 2019), the size of the amplified region, zygosity, clonality, and whether the alteration is truncal (Donoghue et al. 2020).In general, a higher level of amplification may suggest a stronger selective pressure to overexpress a given protein and, therefore, a more appropriate genomic target.Additionally, when multiple genes are involved in a single amplification or deletion event, it can be difficult to know which the tumor was selecting for, as opposed to small amplified or deleted regions that are more likely to be oncogenic (Eifert & Powers 2012, Tanaka & Watanabe 2020).
Zygosity can also influence the likelihood of responding to targeted therapies.In tumor suppressors, biallelic alterations leading to loss of function may be required for therapeutic exploitation.Homozygous alterations, loss of heterozygosity favoring the mutant allele, or epi-genetic silencing of the wild-type allele may indicate functional losses with potential therapeutic relevance (Donoghue et al. 2020).
Truncality/clonality should also be accounted for when selecting therapeutic strategies.Truncal alterations develop early in tumorigenesis and are present across cancer cells.They are likely to be better targets than subclonal alterations in a limited number of cells.Although definitively determining clonality requires complex bioinformatic analysis, the presence of mutations with low mutant allele fractions in comparison to other alterations (accounting for zygosity) or the absence of the alteration in one or more good-quality tumor samples suggests that an alteration may not be clonal.Increasingly, multiregion sequencing and single-cell sequencing can map tumors' clonal architecture.
Although not yet widely available, this technology has the potential to identify patients with intratumoral heterogeneity who are less likely to respond to single-agent targeted therapy, thereby improving patient selection for clinical trials (Dentro et al. 2021, Lim et al. 2020, Liu et al. 2020).

Allele-Specific Differences in Drug Sensitivity
Importantly, different mutations within the same gene may have different susceptibilities to targeted therapy.When these differences are known a priori, they can be accounted for in designing basket trials.In other cases, the trials themselves clarify such biological differences.For example, BRAF alterations are divided into three classes on the basis of how they activate cellular signaling.Currently approved BRAF inhibitors are effective in class I alterations that signal as constitutively active BRAF monomers, in contrast to class II and III alterations that signal as RAF-independent, high-kinase-activity dimers and RAF-dependent, low-kinase-activity dimers, respectively (Fontana & Valeri 2019, Yao et al. 2015).Newer RAF dimer inhibitors are being explored in class I-III BRAF alterations, although inhibition of upstream signaling may be required for effective targeting of class III alterations (Yao et al. 2017(Yao et al. , 2019)).Appreciation of such differences enables rational design of basket trial eligibility criteria.

Histologic Context
The premise of basket trials is that tumors with the same genomic alterations may have shared, exploitable molecular dependencies.It is clear, however, that histology can influence response to targeted therapies.For example, colorectal cancers (CRCs) have demonstrated inferior efficacy compared with other histologies in several basket trials, including those testing BRAF and MEK inhibition (Prahallad et al. 2012, Salama et al. 2020), KRAS G12C inhibition (Fakih et al. 2022, Hong et al. 2020, Ou et al. 2022, Skoulidis et al. 2021), and even TRK inhibition (Drilon et al. 2018).Notably, the tumor-agnostic approval of dabrafenib (a BRAF inhibitor) in combination with trametinib (a MEK inhibitor) for BRAF V600E-mutant cancers explicitly excludes CRCs due to known intrinsic resistance (Gouda & Subbiah 2023).CRC's poor sensitivity to some of these therapies is likely due to feedback activation of EGFR signaling; simultaneous inhibition of EGFR improves outcomes (Amodio et al. 2020, Kopetz et al. 2019, Yaeger et al. 2023).
The utility of poly(ADP-ribose) polymerase (PARP) inhibitors in BRCA1/2-altered cancers similarly illustrates the importance of histologic context.PARP inhibitors exploit lossof-function BRCA mutations using synthetic lethality.Germline BRCA1/2 mutations increase the inherited risk of developing BRCA-associated tumors, namely breast, ovarian, prostate, and pancreatic cancers.Despite the presence of BRCA1/2 alterations across other histologies, the benefit of PARP inhibitors is limited primarily to BRCA-associated tumors, where this alteration appears to be the most biologically relevant (Jonsson et al. 2019, Murciano-Goroff et al. 2022, Schram et al. 2023a).
Histology also has a role in the frequency and mechanisms of acquired resistance.Gastrointestinal cancers harboring TRK fusions are more susceptible to off-target resistance to TRK inhibition mediated by activation of ERK signaling that may be overcome with MAPK pathway inhibition, while other tumor types are more likely to develop on-target TRK mutations that impair drug binding (Cocco et al. 2019).
Eligibility for basket trials must therefore account for available evidence regarding the histologies most likely to benefit, and histologies with predicted resistance may be excluded.For larger basket trials not limited by the rarity of the alteration of interest, individual disease-specific cohorts can be employed and analyzed both individually and in aggregate.

Precision Medicine Knowledge Bases
Online databases have been developed to determine the predictive role of molecular alterations; these include OncoKB, OncoVAR, Jackson Laboratory Clinical Knowledgebase, CIViC, Precision Medicine Knowledgebase, MyCancerGenome, Cancer Genome Interpreter, CancerVar, MetaKB, and ClinVar.OncoKB was the first such database to be designated a public human genetic variant database by the FDA (2021), and it accounts for histologic context and specific alterations in more than 5,000 genes.
Different databases can have disparate interpretations of a variant's oncogenicity (Wagner et al. 2020), and many alterations are still classified as VUSes.While basket trials with inclusive genomic eligibility criteria can help determine the clinical significance of VUSes, their inclusion increases the risk of enrolling patients with tumors not clearly driven by the biomarker of interest.

Challenges in Biomarker Detection
While tissue-based DNA NGS panels that include numerous cancer-associated genes have been widely adopted, they do have limitations.They may detect gene fusions and alternative splice transcripts less reliably and are unable to identify changes in methylation or determine protein expression (Avila & Meric-Bernstam 2019, Di et al. 2019, Schram et al. 2017).RNA-based testing has emerged as a valuable tool to directly detect modified transcripts as well as to uncover targetable fusions and splice variants not detected by DNA testing (Benayed et al. 2019, Schram et al. 2017).This technique is particularly important for genes with large intronic regions that cannot practically be tiled into targeted DNA panels (such as NRG1 fusions) (Nagasaka & Ou 2022).In non-small-cell lung cancer (NSCLC), for example, the integration of RNA and DNA sequencing can detect up to two to three times as many targetable fusions than DNA alone (Harada et al. 2023).An analysis of samples from the multihistology National Cancer Institute Molecular Analysis for Therapy Choice (NCI-MATCH) trial affirmed that many fusions, including actionable alterations with unusual breakpoints and/or unknown partners, may be missed by standard DNA assays (Kaluziak et al. 2023).
The routine incorporation of RNA-based testing into clinical care, while becoming more common, has been limited by the higher cost associated with multiple tests and the fact that RNA is more susceptible to degradation and preanalytical factors than DNA (Harada et al. 2023).Alternative methods also continue to be used.Break-apart fluorescence in situ hybridization is a fast and inexpensive way to detect fusions, but it can test only a limited number of genes simultaneously, cannot identify fusion partners, and can miss fusions with nearby partners or those that are out of frame (Schram et al. 2017).Immunohistochemistry (IHC) is often used to identify biomarkers for trials testing monoclonal antibodies or antibody-drug conjugates (ADCs), but more comprehensive proteomics is required to keep up with the growing list of trial biomarkers.Tissue-based molecular profiling, regardless of the assay used, has the limitation of only testing a sample from a specific site of disease at a single time point, limiting information about spatial and temporal heterogeneity.Liquid biopsies analyzing tumor-derived cellfree DNA (cfDNA) from blood may enable a more comprehensive assessment of tumor heterogeneity and subclones, including resistance alterations, with a shorter turnaround time.Additionally, the ease of blood sampling allows for longitudinal tracking (Heitzer et al. 2019, Keller et al. 2021, Serrano et al. 2020).However, cfDNA poses new challenges for biomarker selection, including the incidental detection of clonal hematopoiesis (CH) and mosaicism.If plasma DNA sequencing alone is used, CH and mosaicism can lead to misinterpretation of mutations as tumor derived and might lead to trial enrollment on the basis of an alteration not present in the clinically relevant tumor (Heitzer et al. 2019, Köhnke & Majeti 2021, Serrano et al. 2020).In a study of patients who underwent paired solid tumor and blood sequencing, at least one CH mutation could be incorrectly attributed to the tumor in roughly 5% of patients (Ptashkin et al. 2018).Another study demonstrated that up to 10% of men with advanced prostate cancer have CH involving DNA repair genes, which have been used as biomarkers for trials in this population (Jensen et al. 2021).Patient-matched normal white blood cell DNA sequencing and bioinformatic algorithms can help distinguish tumor-derived genomic alterations from CH and mosaic variants (Brannon et al. 2021, Serrano et al. 2020).

BASKET TRIALS: PAST, PRESENT, AND FUTURE
Many of the earliest basket trials involved disease-agnostic testing of drug-biomarker pairs that had previously been validated in a single histology in an attempt to expand the population of patients who benefit from targeted therapy.This testing was quickly followed by tumor-agnostic exploration of novel biomarkers.Select biomarkers are described below.

HER2 Overexpression/Amplification and Mutations
HER2 overexpression is a validated biomarker in HER2 + breast and gastric cancer; however, its clinical relevance across tumors is poorly understood.The MyPathway study treated patients with HER2-overexpressed and/or -amplified nonbreast, nongastric cancers with trastuzumab and pertuzumab.Of the 199 KRAS-wild-type, efficacy-evaluable patients in this study, 26% had a confirmed response.Clinical activity differed between histologies, with an objective response rate (ORR) of 64% in salivary cancer; 25-33% in NSCLC, CRC, and biliary and pancreatic cancers; and below 17% in uterine, ovarian, and urothelial cancers (Meric-Bernstam et al. 2021).The national NCI-MATCH program ran a similar study (subprotocol J) of trastuzumab and pertuzumab in HER2-amplified solid tumors.The ORR was 8%, with one response each in colorectal cancer and cholangiocarcinoma (Connolly et al. 2020).A basket trial of the ADC ado-trastuzumab emtansine (T-DM1) similarly demonstrated disease-specific sensitivity, including efficacy in NSCLC and salivary gland carcinoma (Li et al. 2018).Another ADC, trastuzumab-deruxtecan, had an ORR of 61% in HER2 + (3+ by IHC) solid tumors (excluding breast, gastric, and lung cancers).The ORR was 44-85% in cervical, ovarian, endometrial, biliary tract, and bladder cancers, among others (Meric-Bernstam et al. 2023).The drug also has activity in HER2-mutant disease and has recently received approval for previously treated HER2-mutant NSCLC (Li et al. 2022b).HER2 mutations have also been explored pan-cancer.The SUMMIT trial tested the pan-HER kinase inhibitor neratinib in HER2and HER3-mutant tumors.Efficacy varied by both disease type and mutant allele; the greatest activity was observed in breast, cervical, and biliary cancers harboring kinase domain mutations (Hyman et al. 2018).

MAPK Pathway Alterations
The MAPK pathway is commonly altered in cancer and has been the target of many basket trials, including with RAF inhibitors.Vemurafenib was studied in nonmelanoma BRAF V600-mutant tumors, and preliminary activity was identified in NSCLC, Erdheim-Chester disease, and Langerhans cell histiocytosis, among other cancers (Hyman et al. 2015).The international ROAR (Rare Oncology Agnostic Research) and NCI-MATCH subprotocol H trials investigated dabrafenib in combination with trametinib in BRAF V600E-mutant tumors.Activity was observed across disease types (Subbiah et al. 2020(Subbiah et al. , 2022a(Subbiah et al. , 2023a;;Wen et al. 2022), leading to the FDA approval of this combination for solid tumors, excluding CRC (Salama et al. 2020).Several pan-RAF inhibitors and a BRAF-specific dimer breaker are under development for the treatment of BRAF non-V600 mutations with or without the addition of downstream MEK or ERK inhibition (Beck et al. 2023, De La Fuente et al. 2023, Schram et al. 2023b, Sullivan et al. 2020, Wang et al. 2023).
Other MAPK pathway targets have also been tested in basket trials.Initial dose escalation studies of the KRAS G12C inhibitors sotorasib and adagrasib were carried out pancancer, laying the groundwork for their accelerated approval in NSCLC (Hong et al. 2020, Ou et al. 2022).Basket trials are ongoing testing therapy for other KRAS alleles (https://www.clinicaltrials.govidentifiers NCT05379985, NCT05737706, NCT05533463, and NCT05382559, among others).
Inhibitors of downstream and upstream proteins in the MAPK pathway, including MEK, ERK, and SHP2, have been investigated using various MAPK alterations as biomarkers.The efficacy of these inhibitors as monotherapy has generally been limited (Brana et al. 2021, Eckstein et al. 2022, Sullivan et al. 2018), though notable exceptions include MEK inhibition for histiocytosis and low-grade serous ovarian cancer (LGSOC) (Diamond et al. 2019, Gershenson et al. 2022).

PI3K Pathway Alterations
As with the MAPK pathway, several basket trials have targeted the PI3K-AKT pathway.For example, the PIK3CA inhibitor taselisib has been studied across 11 tumor types.While the response rate was limited, activity was observed in select histologies, including cervical as well as head and neck cancers (Jhaveri et al. 2021).
The AKT inhibitor capivasertib has been studied in AKT1 E17K-mutant solid tumors, with responses in cervical, breast, lung, and endometrial cancers (Hyman et al. 2017a).The pan-AKT inhibitor TAS-117 was similarly tested in a basket trial of 13 patients with PI3K/AKT alterations.The ORR was 8%, with activity observed in patients with breast and ovarian cancers harboring select PIK3CA and AKT alterations (Lee et al. 2021).Downstream mTOR inhibition is being explored for patients with alterations in TSC1/2 and/or other PI3K pathway alterations (Burris et al. 2022, Iyer et al. 2023).

FGFR Alterations
Alterations in FGFR1-3, including fusions, amplifications, and mutations, have been observed across tumor types and have formed the basis of basket trials.The inhibitors pemigatinib, futibatinib, and RLY-4008 showed activity in multiple disease types and FGFR alterations (with RLY-4008 being FGFR2-specific), though additional follow-up is needed to determine the utility of targeting FGFR pan-cancer.Efficacy is most consistently demonstrated in FGFR2 fusion-positive intrahepatic cholangiocarcinoma and FGFR3-mutant urothelial cancer (Borad et al. 2023, Meric-Bernstam et al. 2022, Rodon et al. 2023).

Fusions
Oncogenic fusions have served as biomarkers for several notable basket trials.The TRK inhibitor larotrectinib received FDA approval for TRK fusion-positive cancers on the basis of pooled data from three basket trials.In an updated analysis, 244 adult and pediatric patients were efficacy evaluable across 25 different tumor types, most commonly soft tissue sarcoma and thyroid, lung, and salivary gland tumors.Patients had gene fusions involving NTRK1 (46%), NTRK2 (3%), or NTRK3 (51%).The ORR was 69%, with clinically relevant activity observed across diseases and TRK isoforms (Drilon et al. 2022).A pooled analysis from three basket trials (ALKA, STARTRK-1, and STARTRK-2) testing another TRK inhibitor, entrectinib, in TRK fusion-positive cancers demonstrated similar results, with an ORR of 61% (92 of 150 patients) and 17 tumor types represented (Krzakowski et al. 2022).
The entrectinib basket trials also included patients with ROS1 and ALK fusions.In the phase II eligible population, the ORR was 86% among 14 patients with ROS1-rearranged disease (all but one with NSCLC), and an ORR of 57% among seven patients with ALK-rearranged tumors, including NSCLC, renal cell carcinoma, and CRC (Drilon et al. 2017).A similar pediatric trial, the STARTRK-NG trial, enrolled 43 pediatric patients across tumors, with an ORR of 58% (Desai et al. 2022).The NCI-MATCH subprotocols F and G treated patients with ALKor ROS1-rearranged tumors, respectively, with crizotinib.Two of four (50%) eligible ALK fusion-positive patients responded to treatment (one with leiomyosarcoma and the other with CRC).One of four patients with a ROS1 fusion-positive tumor responded (24%), namely a patient with LGSOC (Mansfield et al. 2022).
The RET inhibitor selpercatinib was studied in the phase 1/2 LIBRETTO-001 basket trial for RET fusion-positive tumors.The ORR was 44%.Promising early activity was observed in RET fusion-positive NSCLC as well as in RET mutant-and RET fusion-positive thyroid cancer, leading to FDA approval of selpercatinib in these disease-specific indications followed by tumor-agnostic approval (Drilon et al. 2020, Subbiah et al. 2022b, Wirth et al. 2020).

Microsatellite Instability and High Tumor Mutational Burden
Immunotherapies have received tissue-agnostic approvals based on basket trials.A deficiency in genes responsible for mismatch repair (MMRd) can lead to tumors with genomic microsatellite instability (MSI-H).MMRd/MSI-H tumors are especially susceptible to immune checkpoint blockade (André et al. 2020;Ganesh et al. 2019;Latham et al. 2019;Le et al. 2015Le et al. , 2017)), as are tumors with high tumor mutational burden (TMB), likely due to increased neoantigen production (Samstein et al. 2019).The KEYNOTE-158 trial built on the success of pembrolizumab for MMRd/MSI-H colorectal cancers by testing the drug in other histologies.A total of 27 different MMRd/MSI-H tumor types were represented among 233 patients, with an ORR of 34% (Marabelle et al. 2020b).In a larger analysis of 790 efficacy-evaluable patients for whom TMB testing was available, a 29% response rate was observed among the 102 patients with TMB of 10 or greater, in comparison to 6% among patients with lower TMB (Marabelle et al. 2020a).

DNA Repair
The first reported basket trial enrolled patients with germline BRCA1/2 mutations to the PARP inhibitor olaparib (Fong et al. 2009).BRCA1/2 mutations were enriched in the dose escalation and required for the expansion.As noted above, objective responses were observed only in BRCA1/2 mutation carriers with BRCA-associated cancers.The combination of PARP inhibition and immunotherapy was more recently explored in the JAVELIN BRCA/ATM study, which tested talazoparib along with avelumab, and the MEDIOLA trial, which tested olaparib in combination with durvalumab.These studies also demonstrated notable clinical activity predominantly in BRCA-associated tumor types (Domchek et al. 2020, Krebs et al. 2023, Schram et al. 2023a).Given the success of PARP inhibitors in BRCA-mutant tumors, drugs targeting other proteins important for DNA repair, such as ATR, WEE1, and PKMYT1, are being explored in basket trials using a range of biomarkers (e.g., NCT04855656, NCT04266912, NCT0318896, NCT04855656, NCT04768868).

Novel Targets
Recent basket trials have addressed an expanded range of biologic vulnerabilities of cancer.A growing number of previously undruggable targets have become targetable through advances in chemistry, computer modeling, dynamic simulation, and drug development.KRAS allele-specific inhibitors and multi-inhibitors (Hofmann et al. 2022, Hong et al. 2020, Kemp et al. 2023, Kim et al. 2023, Lito et al. 2016, Ou et al. 2022), p53 Y220C inhibitors (Dumbrava et al. 2022), isoform-selective FGFR inhibitors (Subbiah et al. 2023b), and mutant-specific PIK3CA inhibitors (Perez et al. 2022) all aim to overcome previous drug development challenges.In addition to an expanding list of genetic targets, a growing number of overexpressed surface proteins are being used to direct novel therapies, such as ADCs and bispecific antibodies, to cancer cells (Drago et al. 2021, Meric-Bernstam et al. 2018).Detection of immunologic markers on cancer cells, such as expression of checkpoint proteins or neoantigens (whether a specific neoantigen or assumed neoantigen excess in MSI-H or high-TMB tumors), has enabled basket trials of immune therapies, as discussed above (Friedman et al. 2022, Murciano-Goroff et al. 2020b, Patel et al. 2020, Tateo et al. 2023).
The molecular biomarker for a basket trial no longer needs to be the drug's direct target.Increasingly, basket trials are designed by utilizing knowledge of pathway signaling and synthetic lethality to target the functional consequences of molecular alterations.Examples of attempts to target downstream of the biomarker of interest, which have met with variable success, have included basket trials testing pan-RAF inhibitors and/or MEK and ERK inhibition in RAS-mutant cancers (Chenard-Poirier et al. 2017;Sullivan et al. 2018Sullivan et al. , 2020)); MEK inhibition in NF1-, GNAQ-, and GNA11-mutant tumors (Wisinski et al. 2023); and mTOR inhibitors for the treatment of TSC1/2-, STK11-, PTEN-, and/or PIK3CA-mutant tumors (NCT02465060, NCT04774952).NRG1 fusions are oncogenic ligands that signal through binding to the HER3 receptor.Antibodies targeting HER3 are being explored in tumor-agnostic studies, with promising efficacy reported thus far (Carrizosa et al. 2022, Schram et al. 2022).Similarly, PTCH1 loss-of-function mutations relieve inhibition of SMO, and SMO inhibitors are being investigated in basket trials of PTCH1-altered tumors (NCT02465060).Synthetic lethal approaches being investigated in tumor-agnostic studies include PARP inhibitors for BRCA1/2or ATM-mutant cancers (Schram et al. 2023a); ATR inhibitors for ATM-mutant cancers (Yap et al. 2021); and PKMYT1 inhibition for solid tumors with CCNE1 amplification, FBXW7 loss, and PPP2R1A mutations (NCT04855656).

Novel Therapeutics
To target the growing list of potential biologic vulnerabilities, a broader range of drug classes are being explored beyond traditional small molecules.Basket trials are being designed to test monoclonal and bispecific antibodies, ADCs, proteolysis-targeting chimeras, T cell receptors, and vaccines (Murciano-Goroff et al. 2020a,b).

FACILITATING ENROLLMENT IN BASKET TRIALS
Basket trials entail unique challenges that compound already profound systemic barriers in access to clinical trials.Difficulty identifying appropriate studies, geographic barriers, financial costs, narrow eligibility criteria, attitudes regarding trials, and knowledge of trial availability all limit trial enrollment (Esdaille et al. 2022, Lara et al. 2005, Mohd Noor et al. 2013, Sharrocks et al. 2014, Unger et al. 2021).Fewer than 5% of adult patients with cancer enroll in studies, despite 70% being willing to participate (Comis et al. 2003, Lara et al. 2005, Lynam et al. 2012, Murthy et al. 2004, Tejeda et al. 1996).Enrollment in molecularly selected basket trials has the added barrier of requiring genomic sequencing (which may not be covered by insurance) and appropriate trial matching (Arora et al. 2022, Liu et al. 2022, McCarthy et al. 2016).Additionally, trials for rare genetic alterations may not be available at all centers, creating travel-related financial burdens.Unsurprisingly, patients required to travel more than 120 miles are less likely to enroll in a trial (Uehara et al. 2023).
Recently, the FDA (2022) released a draft guidance outlining measures to facilitate the enrollment of underserved populations through community engagement, language assistance, and reduction of trial burdens (such as allowing the use of local laboratory testing to minimize patient travel).As telemedicine has gained acceptance in the era of COVID-19, many have questioned whether remote care and local monitoring can be further incorporated into trials (Li et al. 2022a).Increased uptake of tumor molecular profiling, expansion of the oncogenic targets being explored, better mechanisms for patient matching, and improved access to trials can maximize equity and efficiency in enrollment.Novel trial designs, such as multicenter master protocols and just-in-time trials, may further facilitate basket trials enrollment.

Next-Generation Sequencing Reports
NGS reports from commercial vendors such as Foundation Medicine, Caris, and Tempus include a nonexhaustive list of potential clinical trials (Simon & Roychowdhury 2013).Some trial sponsors have partnered with genomic sequencing companies to enable direct alerts to providers, though companies have limited clinical information with which to evaluate patient eligibility and dynamic changes in trial availability.

Clinical Trial Matching Services
Some academic centers have created automated trial matching systems.The DARWIN Cohort Management System at Memorial Sloan Kettering Cancer Center sends automated notifications to physicians when patients have been identified who may be eligible for a clinical trial on the basis of a molecular feature (Tao et al. 2019).This matching system is facilitated by a large internal genomic sequencing effort, limiting its generalizability.Similarly, MatchMiner is an open-source platform developed at the Dana-Farber Cancer Institute with the goal of accelerating patient-trial matching (Klein et al. 2022).Several private companies, such as Caris and Foundation Medicine, as well as nonprofit organizations, are developing trial matching services wherein patients provide detailed clinical information, either through engagement with trained trial navigators or through an online interface (see https://www.carislifesciences.com/products-and-services/clinical-trials,https://www.foundationmedicine.com/service/clinical-research-and-trial-matching).

Bringing Trials to the Patient
Another potential solution for facilitating greater trial enrollment is to allow community oncology practices to become on-demand clinical trial sites.In just-in-time trials, local sites undergo activation only when a patient has been identified, eliminating the need for start-up at nonen-rolling centers and allowing smaller sites to provide trial access (Lynam et al. 2012, Wiener et al. 2007).Numerous companies have assisted in this model.For example, TEMPUS has released data showing an average time to activation of 9.4 business days for its trials, far shorter than the estimated baseline average of 20-plus weeks, with a mean time from activation to patient consent of 4.5 days (Oleary et al. 2021).Similarly, the Caris Right-in-Time Clinical Trials program aims for 3 to 12 days to site activation, with patients enrolled by day 14 (see https://www.carislifesciences.com/products-and-services/clinical-trials/right-in-time).

Multicenter Master Protocols
The creation of multicenter, multiarm studies has improved access to basket trials.These studies enroll patients into one of several treatment arms matched to their particular alteration, enabling access to multiple therapies across different trial sites.These master protocols have demonstrated the feasibility of enrolling thousands of patients.The NCI-MATCH and pediatric MATCH trials assigned patients in a disease-agnostic fashion to treatment arms on the basis of molecular alterations (Murciano-Goroff et al. 2021).More than 6,391 patients were enrolled in the adult NCI-MATCH trial, with 18% ultimately to a treatment arm following tumor procurement and sequencing.Response rates were variable, ranging from no responses in certain arms to 38% in the dabrafenib/ trametinib combination arm for BRAF V600E/K-mutant tumors (Murciano-Goroff et al. 2021).The MyPathway and Targeted Agent and Profiling Utilization Registry (TAPUR) trials followed a similar paradigm across countless study sites (Hainsworth et al. 2018, Klute et al. 2022, Mangat et al. 2018, Meric-Bernstam et al. 2019).

Expanded-Access and Single-Patient Protocols
Expanded-access and single-patient protocols provide investigational therapy to patients who cannot enroll in a registrational trial as a result of exclusionary clinical factors or practical barriers (Feit et al. 2019, Scepura et al. 2021).The patient's oncologist is provided with or creates a treatment plan that undergoes institutional review board approval.Efforts to facilitate improved data capture from these patient-centered trials are underway (Polak et al. 2023).

SPECIAL CONSIDERATIONS IN STUDY DESIGN Statistical Considerations
Basket trials pose unique statistical challenges due to the inclusion of heterogeneous diseases with differing bars for clinically relevant efficacy.While a full exploration of basket trial statistics is beyond the scope of this review (Kaizer et al. 2019), it is notable that randomization is rarely feasible given differing standards of care.The majority of basket trials have been exploratory, signal-seeking phase I or II studies that pool results from all histologies and/or report disease-specific efficacy.Alternative two-stage designs have been proposed wherein the inactive indications are pruned at an interim analysis and the active indications are pooled in the final analysis.Given the inflated error potential with cherry-picking, the type 1 error rate needs to be controlled (Chen et al. 2016, Zhou et al. 2019).

Regulatory Considerations
A critical question is what level of evidence justifies disease-agnostic approval versus further testing in individual histologies.Both the US FDA and the European Medicines Agency have offered guidance emphasizing the need for a strong scientific and clinical rationale before considering disease-agnostic approvals (EMA 2021, US Dep. Health Hum. Serv. et al. 2022).More explicit international criteria are likely to emerge, given the growing number of pan-cancer basket trials.

CONCLUSION
Basket trials have become increasingly common in oncology and can be used to achieve tumor-agnostic drug approvals or explore biologic differences in disparate tumor types unified by a common molecular alteration.Several successful genome-directed therapies have stemmed from this study design, providing additional therapeutic options to patients with tumors harboring specific genomic alterations.As the field moves forward, it is critical to optimize biomarker and patient selection, expand molecular testing, streamline clinical trial matching, and improve access to basket studies.

ACKNOWLEDGMENTS
Y.R.M.-G.gratefully acknowledges funding from the Fiona and Stanley Druckenmiller Center for Lung Cancer Research, the Andrew Sabin Family Foundation, and the Society of MSK.A.M.S. gratefully acknowledges funding from a Career Development Award from Conquer Cancer, the ASCO Foundation, a National Cancer Institute (NCI) Cancer Clinical Investigator Team Leadership Award (P30CA008748), and Cycle for Survival.Work on this review was partially supported by an NCI/NIH Cancer Center Support Grant (P30 CA008748) to Memorial Sloan Kettering Cancer Center.DISCLOSURE STATEMENT Y.R.M.-G.reports travel, accommodation, and expenses from AstraZeneca and Loxo Oncology/Eli Lilly.She acknowledges honoraria from Virology Education and Projects in Knowledge (for a continuing medical education program funded by an educational grant from Amgen).She acknowledges associated research funding to the institution from Mirati Therapeutics, Loxo Oncology at Eli Lilly, Elucida Oncology, Taiho Oncology, Hengrui USA, Ltd./Jiangsu Hengrui Pharmaceuticals, Luzsana Biotechnology, Endeavor Biomedicines, and AbbVie.She is an employee of Memorial Sloan Kettering Cancer Center, which has an institutional interest in Elucida.She acknowledges royalties from Rutgers University Press and Wolters Kluwer.She acknowledges food/beverages from Endeavor Biomedicines.Y.R.M.-G.acknowledges receipt of training through an institutional K30 grant (Clinical and Translational Science Award UL1TR00457) from the National Institutes of Health (NIH).She has received funding from a Kristina M. Day Young Investigator Award from Conquer Cancer, the American Society of Clinical Oncology (ASCO) Foundation, endowed by Dr. Charles M. Baum and Carol A. Baum.She is also funded by a Paul Calabresi Career Development Award for Clinical Oncology (NIH/National Cancer Institute K12 CA184746).Y.R.M.-G.acknowledges funding from the Fiona and Stanley Druckenmiller Center for Lung Cancer Research, the Andrew Sabin Family Foundation, and the Society of MSK.G.H. acknowledges honoraria/advisory boards from Lilly, Bristol Myers Squibb, AstraZeneca, MSD, Bayer, and Merck.A.M.S. acknowledges Relay Therapeutics, Mersana (advisory boards); Blueprint Bio, Flagship Pioneering (consulting); Merus, Pfizer (steering committee); and AstraZeneca, ArQule, BeiGene/Springworks, Black Diamond Therapeutics, Elevation Oncology, Kura, Lilly, Merus, Northern Biologics, Pfizer, PMV Pharma, Relay, Repare Therapeutics, Revolution Medicine, and Surface Oncology (research to institution).M.C. reports funding from a Young Investigator Award from Conquer Cancer, the ASCO Foundation, and a Memorial Sloan Kettering Cancer Center T32 Investigational Cancer Therapeutics Training Program grant (T32-CA009207).M.C. holds stock in Nordisk, Quest, Doximity, and Figs.The other authors are not aware of any affiliations, memberships, funding, or financial holdings that might be perceived as affecting the objectivity of this review.LITERATURE CITED Adashek JJ, Subbiah V, Westphalen CB, Naing A, Kato S, Kurzrock R. 2023.Cancer: slaying the nine-headed Hydra.Ann.Oncol 34:61-69 [PubMed: 35931318] Amodio V, Yaeger R, Arcella P, Cancelliere C, Lamba S, et al. 2020.EGFR blockade reverts resistance to KRAS G12C inhibition in colorectal cancer.Cancer Discov.10:1129-39 [PubMed: 32430388] André T, Shiu KK, Kim TW, Jensen BV, Jensen LH, et al. 2020.Pembrolizumab in microsatelliteinstability-high advanced colorectal cancer.N. Engl.J. Med 383:2207-18 [PubMed: 33264544] Arora K, Tran TN, Kemel Y, Mehine M, Liu YL, et al. 2022.Genetic ancestry correlates with somatic differences in a real-world clinical cancer sequencing cohort.Cancer Discov.12:2552-65 [PubMed: 36048199] Avila M, Meric-Bernstam F. 2019.Next-generation sequencing for the general cancer patient.Clin.Adv.Hematol.Oncol 17:447-54 [PubMed: 31449513]

Figure 1 .
Figure 1.Timeline of clinical trials leading to disease-agnostic approvals by the US Food and Drug Administration.Abbreviations: MMRd, mismatch repair deficient; MSI, microsatellite instability; NSCLC, non-small-cell lung cancer; TMB-H, high tumor mutational burden.

Figure 2 .
Figure 2. A schematic showing the relationships between patients, preclinical models, and basket trials that facilitate target validation.Better biomarkers lead to improved patient selection for clinical trials, and basket trials in turn contribute to the biological understanding of biomarkers that can drive future drug development.Figure adapted from images created with BioRender.com.